Cancer prevention by selenium (Se) has been shown to be compound and model dependent, but the mechanisms are not well understood. One hypothesis points to priming of the antioxidant defense system by selenium via modulation of cellular redox parameters and increased expression of antioxidant response element (ARE) genes, including the selenoproteins thioredoxin reductase 1 (TrxRI) and glutathione peroxidase 2 (GPx2). To further investigate the mechanisms of novel and conventional selenocompounds in preventing lung cancer, we propose to test the hypothesis that distinct selenocompounds alter redox status, with ARE/Nrf2 activation and TrxRI and GPx2 playing a role in redox modulation with the following aims: Aim 1: Determine the modulation of redox status of the HOP92 human lung cell line by selenocompounds. This aim focuses on a large cell lung cancer cell line, HOP92, in order to investigate what cellular responses to selenium occur and how these changes compare to the A549 adenocarcinoma and BEAS-2B bronchial epithelial cell lines for which preliminary data exists. Cellular viability will be determined 48 hours after selenocompound treatment using a Cell Counting Kit-8 assay. Cellular redox status will be assessed using cytometric assays for free thiols and ROS. Mechanisms of toxicity will be investigated using a JC-1 cytometric assay for mitochondrial membrane potential and Western blotting for the chaperonin BiP/GRP78. Aim 2: Investigate the role of the Nrf2 transcription factor and selenoproteins TrxRI and GPx2 in modulating oxidative stress induced by selenocompounds in the HOP92, A549 and BEAS-2B cell lines. An ARE reporter construct will be utilized to determine if selenocompound effects are modulated by the Nrf2 transcription factor. Nrf2 activation and requirement will further be validated using mutant ARE reporter constructs and siRNA targeted to Nrf2. Gene knockdowns will also be produced in the three cell lines to reduce transcription of TrxRI and GPx2, which have been found by our lab to be differentially expressed in lung tissue after selenocompound treatment. Cellular responses related to redox status will then be assessed using cytometric assays for free thiols, ROS levels and mitochondrial membrane potential. PUBLIC HEALTH RELEVANCE: Selenium is an essential nutrient present in the diet that has been shown to decrease the incidence and mortality of several types of cancer, including lung cancer. How selenium prevents cancer is currently not well understood, but one way may be by turning on the cell's defense system against oxidative stress. This work seeks to advance the understanding of how selenium supplementation prevents cancer and provide further information on how lung cancer might best be prevented and treated.